Long bones form the major supportive structures of limbs. The ossified tissue first develops around and finally within a cartilage primordium. The maturation of embryonic long bones is dependent on the synchronous transition from cartilage to bone. The long term goal of this project is to understand the cellular regulation of endochondral bone formation. The process may be conveniently studied in the epiphyseal growth plate where the cellular changes occur as a linear continuum. The chondrocytes pass through several stages typified by high mitotic activity or high levels of macromolecular synthesis. Finally, the cells hypertrophy and are replaced by osteogenic cells. Recently, a short collagen molecule, termed type X collagen, has been shown to be a specific product of the hypertrophic chondrocytes. The aims of this study will be to examine the structural function of type X collagen during the formation of endochondral bone. The temporal and spatial distribution of the molecule have been determined by immunohistochemical methods and suggest type X collagen may be involved in the turnover or calcification of the hypertrophic cartilage matrix. The possible role of the molecule in the turnover of the matrix will be assessed by first quantitating the ratio of type X to type II collagen in the matrix. Second, the rates of a mammalian collagenase degradation of types X and II will be compared. Third, hypertrophic cartilage will be analyzed for the presence of enzymes capable of degrading type X collagen. The potential function of the molecule in the calcification of tissue will also be examined. First, the histological distribution of hydroxyapatite will be compared with the location of type X by immunohistochemistry. Second, intramembranous bone will be analyzed biochemically for the presence of type X collagen which has been detected in the tissue by indirect immonufluoresence.